Nop5p Is a Small Nucleolar Ribonucleoprotein Component Required for Pre-18 S rRNA Processing in Yeast*
- From the Department of Anatomy and Cell Biology, Health Science Center, Box 100235, University of Florida, Gainesville, Florida 32610
Abstract
We have identified a novel nucleolar protein, Nop5p, that is essential for growth in Saccharomyces cerevisiae. Monoclonal antibodies B47 and 37C12 recognize Nop5p, which has a predicted size of 57 kDa and possesses a KKXrepeat motif at its carboxyl terminus. Truncations that removed the KKX motif were functional and localized to the nucleolus, but conferred slow growth at 37 °C. Nop5p shows significant sequence homology with yeast Sik1p/Nop56p, and putative homologues in archaebacteria, plants, and human. Depletion of Nop5p in aGAL-NOP5 strain lengthened the doubling time about 5-fold, and selectively reduced steady-state levels of 40 S ribosomal subunits and 18 S rRNA relative to levels of free 60 S subunits and 25 S rRNA. Northern blotting and primer extension analyses showed that Nop5p depletion impairs processing of 35 S pre-rRNA at the A0 and A2 cleavage sites. Nop5p is associated with the small nucleolar RNAs U3, snR13, U14, and U18. Depletion of Nop5p caused the nucleolar protein Nop1p (yeast fibrillarin) to be localized to the nucleus and cytosol. Also, 37C12 co-immunoprecipitated Nop1p. These results suggest that Nop5p functions with Nop1p in the execution of early pre-rRNA processing steps that lead to formation of 18 S rRNA.
Footnotes
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↵* This work was supported by National Institutes of Health Grant GM48586 (to J. P. A) and additional funding for core facility services was provided by the Howard Hughes Medical Institute Research Resources Program of the University of Florida College of Medicine.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
The nucleotide sequence(s) reported in this paper has been submitted to the GenBank™/EMBL Data Bank with accession number(s) AF056070.
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↵‡ To whom correspondence should be addressed. Tel.: 352-392-1873; Fax: 352-392-3305; E-mail: johnaris{at}college.med.ufl.edu.
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↵1 The abbreviations used are: snoRNA, small nucleolar RNA; ETS, externally transcribed spacer; ITS, internally transcribed spacer; mAb, monoclonal antibody; snoRNP, small nucleolar ribonucleoprotein; PAGE, polyacrylamide gel electrophoresis; kb, kilobase(s); 5-FOA, 5-fluoroorotic acid; IP, immunoprecipitation; IF, immunofluorescence.
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↵2 T. Buber and J. P. Aris, unpublished results.
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↵3 V. Lamian and J. P. Aris, unpublished results.
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↵4 S. Chen and J. P. Aris, unpublished results.
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↵5 P. Wei and J. P. Aris, unpublished results.
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↵6 A. C. Metcalf and J. P. Aris, unpublished results.
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- Received January 12, 1998.
- Revision received April 6, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
